This application is a supplement to request salary support for the principal investigator. The scientific portion of the application is little changed from the original. We propose that genetic variation in lysosomal lipases, phospholipases, and diesterases may cause uncharacterized lysosomal storage diseases and may be related to the pathogenesis of atherosclerosis. We have established assay conditons for acid lipase using triglyceride and cholesterol ester substrates and will extend this to include monoglyceride and diglyceride substrates. We have established conditions for acidic phospholipase A1, A2 and C assay in extracts of cultured skin fibroblasts. We have found that these activities are normal in acid lipase deficient cells, but that phospholipase C is deficient in sphingomyelinase deficient cells. Newer plans are to use high pressure liquid chromatography to resolve isozymes of the phospholipases. We also are using (14C) acetate and (32P) phosphate to label phospholipids in cells to demonstrate metabolic alterations. We will perform enzyme purification for acid lipase and selected phospholipases. Measurement of multiple lipase and phospholipase activities on column fractions from enzyme purification will combine with data from mutant cells to further clarify the number of enzymes and genetic loci involved and the specific hydrolytic capacities of each enzyme. We also propose to measure glycerophospholrylcholine diesterase activity as an important unstudied lysosomal function. We will measure phospholipase and diesterase activities in fibroblasts from patients with disorders such as neuronal ceroid lipofuscinosis, sea blue histiocyte disease, "cephalin lipidosis," neuroaxonal dystrophy, and from undiagnosed patients with clinical features of a lysosomal storage disease. Acid lipase will be assayed in lymphocytes of patients undergoing coronary arteriography in an attempt to demonstrate if genetic variation in this enzyme is important in the pathogenesis of atherosclerosis.